Bacteria respond to changes in their chemical and physical environment by appropriately altering their patterns of gene expression or locomotion. The signaling proteins that mediate these sophisticated sensory behaviors must be capable of specifically recognizing one another and of transmitting sensory information through induced conformational changes or reversible covalent modifications. The long-term goal of this work is to understand in molecular terms how bacterial signaling proteins communicate with one another, using the chemotactic behavior of E. coli ad the experimental system. The overall aim of the present proposal is to test the hypothesis that discrete structural domains ("communication modules") promote specific recognition and interaction of bacterial signaling proteins. The experimental plan involves delineation of the structural and functional domains in the CheA, CheW, and CheZ proteins and investigation of the roles of communication modules in promoting signaling transactions among these proteins. Structural domains in the Che proteins will be identified on the basis of resistance to proteolysis and cooperative unfolding at critical denaturant conditions. Genetic approaches will be used to explore the functionally important portions of the proteins by their sensitivity to minor structural changes, and less critical segments of the proteins by their ability to tolerate such mutations. Segments of the che genes will also be isolated and tested for expression of stable polypeptides that inhibit wild type signaling activities to identify putative module coding regions. The in vitro effects of these liberated modules on Che-dependent phosphorylation activities will be compared to their overall effects on chemotactic behavior to explore the signaling functions of interaction domains. Because sensory proteins from a variety of bacteria exhibit common sequence motifs, the lessons learned from the chemotaxis system should be generally applicable to other prokaryotic signaling systems.